Biomonitoring Organochlorine Pesticides in Didelphis virginiana from Yucatan, Mexico by GC-ECD.

The aim of the present work was to apply a validated methodology for the detection of organochlorine pesticides in Didelphis virginiana (Virginia opossum) serum samples collected in Yucatan, Mexico. Recent studies performed to investigate the presence of Organochlorines (OCLs) in water, human blood and milk, and animal tissues from Yucatan have shown that the OCLs concentrations are high and can be associated with potential human health risk. Since opossum is considered an important synanthropic species in Yucatan, 40 opossum serum samples were analyzed by gas chromatography with electron capture detector. The most common OCLs found in opossum sera were lindanes, chlordanes, drines, and endosulfan. Heptachlor, heptachlor epoxide, and lindanes were found at the highest concentrations, while dichlorodiphenyl trichloroethane and its metabolites were found at the lowest concentrations in the samples. The good linearity, precision, and accuracy obtained in the evaluated parameters in the extraction and chromatographic methods support its application for the monitoring of OCLs pesticides in populations of opossums and other wild species in Yucatan.

PHYSIOLOGICAL BASAL PARAMETERS OF FREE-RANGING OPOSSUMS ( DIDELPHIS VIRGINIANA) IN THE YUCATAN PENINSULA, MEXICO.

The aim of the present study is to determine the basal parameters in opossums ( Didelphis virginiana) during physical examination in Yucatan, Mexico. Sixty-six opossums were captured and manually handled for physical examinations. Sex, age, cardiac and respiratory rate, body temperature, pulse, mucous membranes, capillary refill time, hydration, size of superficial lymph nodes, mental status, and body condition were evaluated. Also, comparisons between rectal and auricular temperatures were performed. The means of physiological parameters obtained were: cardiac frequency 146 beats per minute (95% confidence interval [CI]: 138.91-155.24), respiratory frequency 27.33 breaths per minute (95% CI: 25.15-29.64), and body temperature 34.01°C (95% CI: 33.71-34.31). In 54% of animals, a sinus arrhythmia was present. A significant difference was found between rectal and auricular temperature measurements. A variation of -1.64 to 1.14°C was obtained via the auricular thermometer compared with the rectal device.

In situ cytokines (IL-4, IL-10, IL-12, IFN-γ) and chemokines (MCP-1, MIP-1α) gene expression in human Leishmania (Leishmania) Mexicana infection.

Crucial to the defense against Leishmania is the ability of the host to mount a cell-mediated immune response capable of controlling and/or eliminating the parasite. The composition of the cell populations recruited in the early phase of the infection seems to be essential for defining the infection outcomes. The signals that initiate and regulate the early immune response and local accumulation of cell subsets in the skin are poorly understood. We previously studied the in situ expression of cytokine genes in patients with localized cutaneous leishmaniasis (LCL) caused by Leishmania (Leishmania) mexicana. In the present study we examined in situ cytokine (IL-4, IL-10, IL-12, IFN-γ) and chemokine (MCP-1, MIP-1α) gene expression in L. (L.) mexicana active LCL lesions, and in the delayed type hypersensitivity (DTH) skin response to Leishmania antigen in subjects with healed lesion and subclinical infection. Data regarding cytokines were similar to previous studies in patients with active LCL. There were no significant differences in the profile of cytokine and chemokine gene expression in DTH from subjects with healed or subclinical infection. IL-12 gene expression detected in both groups was similar. High expression of MCP-1 was detected in all patients with active LCL. There was no difference in the level of MCP-1 expression between the healed lesion and the subclinical infection groups (p = 0.876). IL-12 and MCP-1 in the absence of IFN-γ might be playing a crucial role in infection outcomes at skin level.

Spatial distribution of Triatoma dimidiata peridomestic colonies modulated by distance between susceptible microhabitat patches.

Triatoma dimidiata is a vector of the hemoparasite Trypanosoma cruzi, the causal agent of Chagas disease. It settles reproductive colonies in the peridomicile of the premises. The peridomicile is comprised of a random set of artificial and natural features that overlap and assemble a network of microenvironmental suitable sites (patches) that interact with each other and favor the structure and proliferation of T. dimidiata colonies. The heterogeneity of patch characteristics hinders the understanding and identification of sites susceptible to colonization. In this study, a classification system using a random forest algorithm was used to identify peridomiciles susceptible to colonization to describe the spatial distribution of these sites and their relationship with the colonies of T. dimidiata in ten localities of Yucatan. From 1,000 peridomiciles reviewed, the classification showed that 13.9 % (139) of the patches were highly susceptible (HSP), and 86.1 % (861) were less susceptible (LSP). All localities had at least one HSP. The occupancy by patch type showed that the percentage of total occupancy and by colonies was higher in the HSP, while the occupancy by adult T. dimidiata without evidence of nymphs or exuviae (propagules) was higher in the LSP. A generalized additive model (GAM) revealed that the percentage of occupied patches increases as the abundance of individuals in the localities increases however, the percentage of occupied patches in LSP is lower than occupied in HSP. Distance analyses revealed that colonies and propagules were located significantly closer (approximately 200 m) to a colony in a HSP than any colony in a LSP. The distribution of T. dimidiata in the localities was defined by the distribution of patch type; as the occupancy in these patches increased, a network of peridomestic populations was configured, which may be promoted by a greater abundance of insects inside the localities. These results reveal that the spatial distribution of T. dimidiata individuals and colonies in the peridomicile at the locality scale corresponds to a metapopulation pattern within the localities through a system of patches mediated by distance and level of the vectors' occupancy.

Seasonal and pandemic influenza H1N1 viruses induce differential expression of SOCS-1 and RIG-I genes and cytokine/chemokine production in macrophages.

BACKGROUND: Infection with pandemic (pdm) A/H1N1 virus induces high levels of pro-inflammatory mediators in blood and lungs of experimental animals and humans. METHODS: To compare the involvement of seasonal A/PR/8/34 and pdm A/H1N1 virus strains in the regulation of inflammatory responses, we analyzed the changes in the whole-genome expression induced by these strains in macrophages and A549 epithelial cells. We also focused on the functional implications (cytokine production) of the differential induction of suppressors of cytokine signaling (SOCS)-1, SOCS-3, retinoid-inducible gene (RIG)-I and interferon receptor 1 (IFNAR1) genes by these viral strains in early stages of the infection. RESULTS: We identified 130 genes differentially expressed by pdm A/H1N1 and A/PR/8/34 infections in macrophages. mRNA levels of SOCS-1 and RIG-I were up-regulated in macrophages infected with the A/PR/8/34 but not with pdm A/H1N1 virus. mRNA levels of SOCS-3 and IFNAR1 induced by A/PR/8/34 and pdm A/H1N1 strains in macrophages, as well as in A549 cells were similar. We found higher levels of IL-6, TNF-α, IL-10, CCL3, CCL5, CCL4 and CXCL8 (p < 0.05) in supernatants from cultures of macrophages infected with the pdm A/H1N1 virus compared to those infected with the A/PR/8/34 strain, coincident with the lack of SOCS-1 and RIG-I expression. In contrast, levels of INF-α were higher in cultures of macrophages 48h after infection with the A/PR/8/34 strain than with the pdm A/H1N1 virus. CONCLUSIONS: These findings suggest that factors inherent to the pdm A/H1N1 viral strain may increase the production of inflammatory mediators by inhibiting SOCS-1 and modifying the expression of antiviral immunity-related genes, including RIG-I, in human macrophages.

Exposure to Trypanosoma cruzi and Leishmania parasites in dogs from a rural locality of Yucatan, Mexico. A serological survey.

Trypanosoma cruzi, the causal agent of American trypanosomiasis, and Leishmania spp., the causal agents of Leishmaniasis, are prevalent in more than 20 American countries, including Mexico. Dogs have been reported as incidental hosts for both parasites and may be helpful as transmission sentinels. We surveyed the dog population in a rural locality of the Merida municipality in Yucatan, Mexico, to evaluate the seroreactivity against T. cruzi and Leishmania spp. using two antigens, parasite homogenate (H) and iron superoxide dismutase extract (FeSODe), with two serological techniques (ELISA and Western Blot). Our study found that 3.33% of the tested dogs were seroreactive to T. cruzi using ELISA-H, and 29.5% were seroreactive to FeSODe antigen, with a 94.4% consistency between the two tests. Similarly, for L. mexicana, 1.6% were seroreactive using ELISA-H, and 9.8% were seroreactive using ELISA-FeSODe, with an 83.3% consistency between tests. For L. braziliensis, no dogs were seroreactive using ELISA-H, but 16.4% were seroreactive using ELISA-FeSODe, with a 90% consistency between tests. Finally, for L. infantum, 4.9% were seropositive using ELISA-H, and 6.6% were seropositive using ELISA-FeSODe, with a 75% consistency between tests. These results show noticeable evidence of exposure of dogs to trypanosomatid parasites and highlight the potential disease risk for the people and their companion animals in the region.

Molecular Detection of Leishmania (Leishmania) mexicana in Sandflies from the State of Yucatan, Mexico.

Background: Leishmaniases are a group of vector-borne zoonotic diseases of public health relevance within the tropical and subtropical regions of the world. The state of Yucatan is a vulnerable and receptive area to localized cutaneous leishmaniasis (LCL) due to its proximity to the high-transmission endemic states of Campeche and Quintana Roo. Autochthonous cases of LCL caused by Leishmania (Leishmania) mexicana have been documented in the state, showing a geographical expansion of the disease. Materials and Methods: Using CO2-supplemented Centers for Disease Control and Prevention light traps and Shannon traps, we captured anthropophilic sandflies in the surroundings of a locality with recent records of autochthonous cases of LCL. Sandflies carrying Leishmania DNA were evidenced using PCR. Results: A total of 140 Phlebotominae (Diptera: Psychodidae) females of four species were captured: Lutzomyia (Tricholateralis) cruciata (Coquillett), Psathyromyia (Psathyromyia) shannoni (Dyar), Lutzomyia (Lutzomyia) longipalpis (Lutz and Neiva), and Dampfomyia (Coromyia) deleoni (Fairchild and Hertig). Molecular results showed that 6.1% (95% confidence interval [CI] = 2.3-12.9%) of Lu. cruciata and 43.8% (95% CI = 19.8-70.1%) of Pa. shannoni showed evidence of carrying L. (L.) mexicana DNA. Conclusion: We provide evidence of anthropophilic sandflies carrying L. mexicana DNA in a municipality with recorded autochthonous cases of LCL caused by this parasite species in the state of Yucatan, suggesting the emergence of new focus of LCL in Mexico.

Trypanosoma cruzi and Leishmania spp. in rodents in a peri-urban area of Central Mexico.

Rodents are one of the most relevant groups of mammals involved in the process of zoonotic disease transmission. Their ability to adapt to anthropized environments allows them to come into contact with humans with often negative consequences for the latter. The present study designed to detect the presence of Trypanosoma cruzi and Leishmania spp. in rodents living in the peri-urban area of Queretaro in central Mexico. This research was carried out during two seasons of collection of wild and domestic rodents, in three localities within the peri-urban area of the state of Queretaro. These collections were carried out during the dry season of February-May 2017 and in the rainy season of August-November 2017. Samples were obtained from the tail tip, from which DNA was purified using the DNeasy Blood & Tissue Kit. End-point PCR was used for the identification of Trypanosoma cruzi and Leishmania spp. A total of 82 rodents were caught, represented in three families, six genera and seven species, of which 29 (35.3%) were positive for Trypanosoma cruzi; 13 (15.8%) for Leishmania spp.; and 12 individuals presented co-infection with both parasites (14.6%). This study confirmed the presence of Trypanosoma cruzi and Leishmania spp. in synanthropic rodents in the peri-urban area of Queretaro, where Chagas and Leishmaniosis diseases are not considered endemic. It is necessary to continue researching for the presence of vectors, as well as for the detection of diseases caused by parasites in humans and thus be able to confirm the transmission cycle of Trypanosoma cruzi and Leishmania spp. in this central Mexican city.

Cutaneous Leishmaniasis Emergence in Southeastern Mexico: The Case of the State of Yucatan.

Environmental changes triggered by deforestation, urban expansion and climate change are present-day drivers of the emergence and reemergence of leishmaniasis. This review describes the current epidemiological scenario and the feasible influence of environmental changes on disease occurrence in the state of Yucatan, Mexico. Relevant literature was accessed through different databases, including PubMed, Scopus, Google, and Mexican official morbidity databases. Recent LCL autochthonous cases, potential vector sandflies and mammal hosts/reservoirs also have been reported in several localities of Yucatan without previous historical records of the disease. The impact of deforestation, urban expansion and projections on climate change have been documented. The current evidence of the relationships between the components of the transmission cycle, the disease occurrence, and the environmental changes on the leishmaniasis emergence in the state shows the need for strength and an update to the intervention and control strategies through a One Health perspective.

Report of autochthonous cases of localized cutaneous leishmaniasis caused by Leishmania (Leishmania) mexicana in vulnerable, susceptible areas of Southeastern Mexico.

Localized cutaneous leishmaniasis (LCL) is an endemic disease in several Mexican States with the main endemic areas located in the South-Southeast region of the country, where 90% of Leishmania (Leishmania) mexicana cases are registered. The Southeast region is located in the Yucatan Peninsula, including Campeche, Quintana Roo and Yucatan States. Campeche and Quintana Roo register more than 60% of the cases in the country each year, while in Yucatan the reports are of imported cases due to residents traveling to endemic areas. However, since 2015, autochthonous cases have been diagnosed by health authorities in municipalities with no previous transmission records. We aimed to identify Leishmania parasite species involved in autochthonous cases by means of the PCR technique. The present study included 13 autochthonous cases of LCL with clinical and parasitological diagnoses during 2018 and 2019 by health authorities, without specific identification of the causal agent. Tissue samples were taken by scraping the margins of active lesions and then they were spotted onto an FTATM Elute Microcard. Next, DNA was eluted and used for PCR amplification of specific Leishmania genus and L. (L.) mexicana species-specific fragments. Molecular analysis showed evidence that L. (L.) mexicana was the causal agent of LCL in 12 of the 13 patients; in one patient, PCR was not performed due to the patient's refusal to participate in the study. Identifying Leishmania species that cause LCL is necessary to define efficient treatment schemes and control strategies for the disease in vulnerable and susceptible areas of the Yucatan State's municipalities.

CRTAM: A molecule involved in epithelial cell adhesion.

Class I-restricted T cell associated molecule (CRTAM) is a member of the immunoglobulin superfamily that complies with the structural characteristics of the JAM family of proteins and is phylogenetically more closely related to nectin-like proteins. Here we demonstrate for the first time, that CRTAM is expressed in epithelial cells along the lateral membrane and is important for early cell-cell contacts and cell-substrate interactions. CRTAM is sensitive to intermediate filament disruption and treatment of monolayers with soluble CRTAM enhances cell-cell dissociation and lowers transepithelial electrical resistance. Incubation of newly plated cells with anti-CRTAM antibody decreases the formation of cell aggregates and promotes cell detachment. Co-cultures of epithelial cells and fibroblasts that lack CRTAM expression and in vitro binding assays, demonstrate the participation of CRTAM in homotypic and heterotypic trans-interactions. Hence we conclude that CRTAM is a molecule involved in epithelial cell adhesion.

Class I-Restricted T Cell-Associated Molecule Is a Marker for IFN-γ-Producing iNKT Cells in Healthy Subjects and Patients with Type 1 Diabetes.

Class I-restricted T cell-associated molecule (CRTAM) is an activation marker expressed on the cell surface of activated invariant natural killer T (iNKT) cells, CD8+ T cells, and a small subset of CD4+ T cells. CRTAM has also been associated with a proinflammatory profile in murine CD4+ T cells. However, CRTAM has not been thoroughly explored in human cells. This work focused on evaluating CRTAM expression in human iNKT lymphocytes after activation with α-galactosylceramide, its widely used specific glycolipid antigen. We also analyzed the involvement of costimulatory molecules in CRTAM expression and whether CRTAM expression is associated with a specific effector cytokine profile. We found that the signal produced by invariant T cell receptor (iTCR) engagement with α-galactosylceramide is sufficient to trigger CRTAM expression on human iNKT cells after 18 h of stimulation. Moreover, we observed a clear association between CRTAM expression and IFN-γ production in iNKT cells from healthy subjects and patients with type 1 diabetes. However, blocking the engagement of costimulatory molecules, such as CD40, CD80, and CD86, did not modify CRTAM expression. These results indicate that CRTAM may also play a role in triggering the production of IFN-γ in human iNKT cells and that CRTAM could be used as a marker to identify these inflammatory cells.

Report of autochthonous cases of localized cutaneous leishmaniasis caused by Leishmania (Leishmania) mexicana in vulnerable, susceptible areas of Southeastern Mexico

ABSTRACT Localized cutaneous leishmaniasis (LCL) is an endemic disease in several Mexican States with the main endemic areas located in the South-Southeast region of the country, where 90% of Leishmania (Leishmania) mexicana cases are registered. The Southeast region is located in the Yucatan Peninsula, including Campeche, Quintana Roo and Yucatan States. Campeche and Quintana Roo register more than 60% of the cases in the country each year, while in Yucatan the reports are of imported cases due to residents traveling to endemic areas. However, since 2015, autochthonous cases have been diagnosed by health authorities in municipalities with no previous transmission records. We aimed to identify Leishmania parasite species involved in autochthonous cases by means of the PCR technique. The present study included 13 autochthonous cases of LCL with clinical and parasitological diagnoses during 2018 and 2019 by health authorities, without specific identification of the causal agent. Tissue samples were taken by scraping the margins of active lesions and then they were spotted onto an FTATM Elute Microcard. Next, DNA was eluted and used for PCR amplification of specific Leishmania genus and L. (L.) mexicana species-specific fragments. Molecular analysis showed evidence that L. (L.) mexicana was the causal agent of LCL in 12 of the 13 patients; in one patient, PCR was not performed due to the patient's refusal to participate in the study. Identifying Leishmania species that cause LCL is necessary to define efficient treatment schemes and control strategies for the disease in vulnerable and susceptible areas of the Yucatan State's municipalities.

Reduced iNKT cells numbers in type 1 diabetes patients and their first-degree relatives.

Type 1 diabetes (T1D) is an autoimmune disease that is characterized by the specific destruction of insulin-producing pancreatic ß cells. Invariant natural killer T (iNKT) cells have been associated with development of T1D. Class I MHC-restricted T cell-associated molecule (CRTAM) is expressed on activated iNKT, CD8(+), and CD4(+) T cells, and it is associated with the pro-inflammatory profiles of these cells. Crtam gene expression in CD3(+) lymphocytes from non-obese diabetic (NOD) mice is associated with T1D onset. However, expression of CRTAM on T cells from patients with T1D has not yet been evaluated. We compared iNKT cell (CD3(+)Vα24(+)Vß11(+)) numbers and CRTAM expression in a Mexican population with recent-onset T1D and their first-degree relatives with control families. Remarkably, we found lower iNKT cell numbers in T1D families, and we identified two iNKT cell populations in some of the families. One iNKT cell population expressed high iTCR levels (iNKT(hi)), whereas another expressed low levels (iNKT(lo)) and also expressed CRTAM. These findings support a probable genetic determinant of iNKT cell numbers and a possible role for these cells in T1D development. This study also suggests that CRTAM identifies recently activated iNKT lymphocytes.

The role of natural killer T (NKT) cells in the pathogenesis of type 1 diabetes.

In type 1 diabetes, a failure in the regulation of either innate or acquired immunity may be the cause of autoimmune response. A cell population that may have a regulatory role of the immune response are the Natural Killer T (NKT) cells, which are a population expressing T lymphocyte antigen receptor (TCR), and are a common marker for NK cells. A distinctive characteristic in NKT cells is their capacity to produce large amounts of immune-modulating cytokines. A decrease in the number and/or functional incapability of NKT cells is associated with progression of type 1 diabetes and with other self-immune diseases. However, the relevance of such findings is not completely understood. Limitations of the current studies include the existing methods to measure NKT activation and the lack of assessment of the expression of genes affected by NKT action. Nevertheless, the study of NKT cells may be a new clinical approach to detect individuals at risk for having type 1 diabetes. Additional studies are needed to evaluate the clinical value of this new predictive tool.

Role of CRTAM during mouse early T lymphocytes development.

CRTAM was reported as a novel receptor expressed in activated NKT and CD8 T lymphocytes. However, we have recently shown that it is also expressed in several non-immune tissues. In opposition to what has been stated for lymphoid cells, CRTAM expression is constitutive in epithelia, suggesting a role in cell-cell interactions. Given the importance of cell interactions during T lymphocyte development, we evaluated CRTAM during T lymphocyte ontogeny. Here we show that CRTAM has an unexpected constitutive expression in adult thymocytes and, remarkably, it is sustained during all stages of thymocyte development. CRTAM expression is restricted to CD8 and all DN subpopulations, with a consistent pattern from E13.5 stage to adult mice. Blocking CRTAM interaction with CADM1 impairs thymus growth, uncovering a novel role in thymus development, with a consequent impact in thymocyte maturation. Thus, CRTAM interaction with CADM1 is involved in structural maintenance of the thymic lobes.

Characterization of CRTAM gene promoter: AP-1 transcription factor control its expression in human T CD8 lymphocytes.

Class-I MHC-restricted T-cell associated molecule (CRTAM) is a member of the Nectin-like adhesion molecule family. It is rapidly induced in NK, NKT and CD8(+) T cells. Interaction with its ligand Nectin-like 2 results in increased secretion of IFN-gamma by activated CD8(+) T lymphocytes. Through sequential bioinformatic analyses of the upstream region of the human CRTAM gene, we detected cis-elements potentially important for CRTAM gene transcription. Analyzing 2kb upstream from the ATG translation codon by mutation analysis in conjunction with luciferase reporter assays, electrophoretic mobility shify assay (EMSA) and supershift assays, we identified an AP-1 binding site, located at 1.4kb from the ATG translation codon of CRTAM gene as an essential element for CRTAM expression in activated but not resting human CD8(+) T cells. CRTAM expression was reduced in activated CD8(+) T cells treated with the JNK inhibitor SP600125, indicating that CRTAM expression is driven by the JNK-AP-1 signaling pathway. This study represents the first CRTAM gene promoter analysis in human T cells and indicates that AP-1 is a positive transcriptional regulator of this gene, a likely important finding because CRTAM has recently been shown to play a role in IFN-gamma and IL-17 production and T cell proliferation.